While vaccination programs have clear documented success in controlling many diseases, there has been a failure to generate effective, long-term immunity against certain major pathogens. On the other hand, in carcinogenic situations there is an urgent need to develop therapies that promote the host immune system to target and destroy cancerous tumors and metastases. Mesenchymal stem cells (MSC) are unique multipotent progenitor cells that are presently being exploited as gene therapy vectors for a variety of conditions, including cancer and autoimmune diseases (Klopp et al., 2007; Le Blanc and Ringden, 2007; Spaeth et al., 2008; Bergfeld and DeClerck, 2010; Chen et al., 2010; Liang et al., 2010; Martino et al., 2010; Panes et al., 2010). Although MSC are predominantly known for anti-inflammatory properties during allogeneic MSC transplant, there is evidence that MSC can actually promote adaptive immunity under certain settings. MSC have been identified in a wide variety of tissues, including bone marrow, adipose tissue, placenta, and umbilical cord blood. Adipose tissue is one of the richest known sources of MSC.
MSC have been successfully transplanted into allogeneic hosts in a variety of clinical and pre-clinical settings (Di Nicola et al., 2002; Meisel et al., 2004; Aggarwal and Pittenger, 2005; Chen et al., 2006; Corcione et al., 2006; Sotiropoulou et al., 2006; Uccelli et al., 2007; Raffaghello et al., 2008). These donor MSC often promote immunotolerance (Potian et al., 2003; Aggarwal and Pittenger, 2005), including the inhibition of graft-versus-host disease (GvHD) that can develop after cell or tissue transplantation from a major histocompatibility complex (MHC)-mismatched donor (Ringden et al., 2006; Wernicke et al., 2011). The diminished GvHD symptoms after MSC transfer has been due to direct MSC inhibition of T and B cell proliferation, resting natural killer cell cytotoxicity, and DC maturation (reviewed in (Uccelli et al., 2008)). At least one study has reported generation of antibodies against transplanted allogeneic MSC (Sundin et al., 2007). Nevertheless, the ability to prevent GvHD also suggests that MSC expressing foreign antigen might have an advantage over other cell types (i.e., dendritic cells; DC) during a cellular vaccination in selectively inducing immune responses to only the foreign antigen(s) expressed by MSC and not specifically the donor MSC. Use of MSC as the cellular base for an alternative vaccination strategy may save on production time and costs associated with necessary HLA matching if other cell types were used.
The use of modified MSC also is being explored in vivo in order to enhance the immunomodulatory properties of MSC (Choi et al., 2008; Sasaki et al., 2009; Kumar et al., 2010; Klinge et al., 2011). MSC transduced to overproduce IL-10 suppressed collagen-induced arthritis in a mouse model (Choi et al., 2008). In addition, MSC expressing glucagon-like peptide-1 transplanted into an Alzheimer's disease mouse model led to a decrease in A-beta deposition in the brain (Klinge et al., 2011). In an osteopenia mouse model, mice receiving transduced MSC that had stable expression of bone morphogenetic protein had increased bone density (Kumar et al., 2010). In a rat model for spinal cord injury, rats treated with MSC stably overexpressing of brain-derived neurotrophic factor had a better overall outcome than rats administered MSC alone (Sasaki et al., 2009). Lastly, in a rat model for bladder outlet obstruction, rats receiving transduced MSC with stable overexpression of hepatocyte growth factor had decreased collagen accumulation in the bladder (Song et al., 2012). These studies indicate that modified MSC are a useful and feasible vehicle for protein expression/delivery to target various diseases and tissues.
MSC have been studied as a delivery vehicle for anti-cancer therapeutics due to their innate tendency to home to tumor microenvironments, and is thoroughly reviewed in (Loebinger and Janes, 2010). MSC have also been used to promote apoptosis of tumorigenic cells through the expression of IFNα or IFNγ (Li et al., 2006; Ren et al., 2008). Additionally, MSC have recently been explored for the prevention and inhibition of tumorigenesis and metastasis. A study by Wei et al. examined the use of human papilloma virus (HPV)-immortalized MSC that express the HPV proteins E6/E7 combined with a modified E7 fusion protein vaccine in a mouse tumor model where metastatic fibrosarcoma cells were administered (Wei et al., 2011). This group found that only mice that were immunized with both the E7-expressing MSC and modified E7 protein vaccine showed a decrease in tumor growth, and an E7-specific antibody response. Mice receiving either MSC or protein vaccine alone were not able to raise an anti-E7 response or inhibit tumor growth of metastatic sarcoma. The limitation of this interesting approach is that it can only be used as an anti-cancer therapeutic and not as a universal cancer preventative, as individual tumors have unique antigen expression. In addition, a long-term safety examination of these immortalized MSC/protein vaccine therapy in cancer-free mice is warranted. Although these immortalized MSC were previously determined to be non-tumorigenic (Hung et al., 2004), they persisted in mice longer than 21 days, unlike primary MSC (i.e. non-immortalized), which are only detectable for a very short time after administration (Gao et al., 2001; Abraham et al., 2004; Ohtaki et al., 2008; Prockop, 2009). Thus, there may be unforeseen outcomes in the long term (i.e. outcompeting with endogenous MSC and differing immunomodulatory abilities, which were not assessed in this study) with the use of immortalized MSC even if they prove to be non-malignant. Other studies have indicated that immortalized MSC can become tumorigenic, and thus must be carefully studied to determine if they are indeed safe for use (Rubio et al., 2005; Phinney and Prockop, 2007; Tolar et al., 2007).
Vaccines generally are considered to be one of the most efficient and cost-effective means of preventing infectious disease. Vaccines have already demonstrated transformative potential in eradicating one devastating disease, smallpox, while offering the ability to control other diseases, including diphtheria, polio, and measles, that formerly caused widespread morbidity and mortality. The development of vaccines involves the testing of an attenuated or inactivated version of the pathogen or identification of a pathogen component(s) (i.e. subunit, toxoid, virus-like particle) that elicits an immune response that protects recipients from disease when they are exposed to the actual pathogen. In an ideal world a single vaccine would be able to target all major human pathogens (versatile), elicit strong protective immunity to these pathogens (robust) without inducing unwanted side-effects (safe), and still be fairly inexpensive to produce per dose (cost-effective). In the case of viruses or host-cell produced proteins, vaccine production that includes human post-translational processing, mimicking natural infection, will likely prove to be superior to bacterial or other expression systems.
Traditional vaccine approaches have thus far failed to provide protection against HIV, tuberculosis, malaria and many other diseases, including dengue, herpes and even the common cold. The reasons why traditional vaccine approaches have not been successful for these diseases are complex and varied. For example, HIV integrates functional proviral genomes into the DNA of host cells, thereby establishing latency or persistence. Once latency/persistence is established, it has not been possible to eradicate HIV, even with highly active antiretroviral therapy. Clearly, new approaches to vaccine development are needed to address HIV and other intractable vaccine challenges.
Newer alternative immunization approaches include both DNA and cellular vaccines. DNA vaccines involve the transfection of cells at the tissue site of vaccination with an antigen-encoding plasmid that allows local cells (i.e. myocytes) to produce the vaccine antigen in situ. Cellular vaccines use the direct transfer of pre-pulsed or transfected host cells (i.e. dendritic cells, DC) expressing or presenting the vaccine antigen. The advantage of these approaches is that vaccine antigens are produced in vivo and are readily available for immunological processing. Despite numerous reports of successful pre-clinical testing, both such approaches have hit stumbling blocks. DNA vaccination studies in humans show poor efficacy, which was linked to innate differences between mice and humans (Cavenaugh et al., 2011; Wang et al., 2011). DC vaccination strategies have shown limited clinical success for therapeutic cancer vaccinations and have high production costs due to necessary individual tailoring (Bhargava et al., 2012; Palucka and Banchereau, 2012).
There is an ongoing need for new strategies for vaccination against infectious diseases. The episomally transfected MSC and methods described herein address these needs.